Intracellular free calcium and intracellular pH play a critical role in cell proliferation, differentiation and death. The Ca+ regulating hormone 1,25(OH)2-vitamin D3 (1,25(OH)2D3) inhibits proliferation of breast cancer cells, but its therapeutic potential is limited by hypercalcemia. The antiproliferative activity of 1,25(OH)2D3 analogues (deltanoids) does not depend significantly on vitamin D receptors (VDR) present in breast cancer cells. Nongenomic effects of 1,25(OH)2D3, particularly on calcium and pH, may determine, to a substantial extent, its role in cellular homeostasis. The authors have shown that several analogs induce non- genomic versus genomic effects, and that 1,25(OH)2D3 regulates calcium entry into the cell through voltage-insensitive calcium channels (VICC) coupled to plasmalemmal VDR, and mobilizes the ryandone-sensitive endoplasmic reticulum calcium stores. The purpose of this project is to determine the mechanism by which 1,25(OH)2D3 exerts its antiproliferative effects. This will be done by testing whether the D3 analog-agonists increase Ca+ influx, causing apoptosis, or whether analog-antagonists inhibit calcium entry and suppress cell proliferation. Comparison of these effects, along with prodifferentiating activities of the analogs versus binding to VDR, the ability of the analogs to activate VICC and mobilize calcium stores, will help in understanding the roles of D3 and calcium in breast cancer, and will be useful in designing new anticancer d3 analogs.